This invention is concerned with recombinant DNA molecules, and methods for producing proteins by said molecules. The invention is particularly concerned with recombinant DNA molecules that are synthesized in Bacillus strain bacteria and are known to have DNA which codes for exoenzymes excreted by them and that are present in tens of copies in Bacillus strain bacteria; as well as with recombinant DNA molecules modified from the above recombinant DNA molecules that are known to have DNA which contains the regulation and excretion signals of the .alpha.-amylase gene of B. amyloliquefaciens, to which signals a gene of any protein can be joined. As will be described in the following, these recombinant DNA molecules can be used, for example, to intensify the .alpha.-amylase production in Bacillus strain bacteria, and their modifications to produce any protein in Bacillus strain bacteria.
Resent development in molecular biology has created new possibilities for protein production in bacteria by recombinant DNA techniques. In addition to the possibility of producing proteins of eukaryotic cells in bacteria by recombinant DNA techniques, the synthesis of the proteins of the bacteria themselves can be significantly improved by increasing the number of the copies of the desired gene in the cell. The number of the gene copies in a bacterium cell can be increased by joining the gene to such a plasmid or virus DNA molecule as is found in the cell in several, usually 10 to 100, copies. The increased number of the gene copies in a cell usually also leads to a corresponding increase in the protein synthesis expressed by the gene.
Even though several experiments of this type have been carried out using E. coli and plasmid or virus DNA molecules replicating in it as host bacterium, the use of Bacillus strain bacteria as hosts is only beginning (Bryczan et al., Molecular General Genet. 177, 459-467, 1979; Keggins et al., Proc. Natl. Acad. Sci. U.S.A. 75, 1423-1427, 1978; Yoneda et al., Biochem. Biophys. Res. Commun., 91, 1556-1564, 1979). None of the methods publicised so far are concerned with increasing the production of the exoenzyme of a Bacillus strain bacterium in Bacillus strain bacteria in a manner which would allow the gene coding for the exoenzyme to be replicated by joining it to the plasmid that is present in the Bacillus strain bacterium in several copies (Part I of the invention), nor are any of the publicised methods concerned with producing proteins by a method in which the regulation and secretion signals of the gene of the enzyme secreted by the Bacillus strain bacteria have been joined to the gene of the protein desired to be produced (Part II of the invention). As an example of the 1st part of the invention, by which the production of Bacillus strain bacterium exoenzymes can be intensified through increasing the number of the genes of the desired exoenzyme in the cell, the transfer of the Bacillus-.alpha.-amylase gene is presented.